Combined effects of polyamide microplastic and sulfamethoxazole in modulating the growth and transcriptome profile of hydroponically grown rice (Oryza sativa L.).

The use of reclaimed water from wastewater treatment plants for irrigation has a risk of introducing micropollutants such as microplastics (MPs) and antimicrobials (AMs) into the agroecosystem. This study was conducted to investigate the effects of single and combined treatment of 0.1 % polyamide (PA ∼15 µm), and varying sulfamethoxazole (SMX) levels 0, 10, 50, and 150 mg/L on rice seedlings (Oryza sativa L.) for 12 days. The study aimed to assess the impact of these contaminants on the morphological, physiological, and biochemical parameters of the rice plants. The findings revealed that rice seedlings were not sensitive to PA alone. However, SMX alone or in combination with PA, significantly inhibited shoot and root growth, total biomass, and affected photosynthetic pigments. Higher concentrations of SMX increased antioxidant enzyme activity, indicating oxidative stress. The roots had a higher SMX content than the shoots, and the concentration of minerals such as iron, copper, and magnesium were reduced in roots treated with SMX. RNA-seq analysis showed changes in the expression of genes related to stress, metabolism, and transport in response to the micropollutants. Overall, this study provides valuable insights on the combined impacts of MPs and AMs on food crops, the environment, and human health in future risk assessments and management strategies in using reclaimed water.

Impact of geochemistry and microbes on the methylmercury production in mangrove sediments.

Unraveling the geochemical and microbial controls on methylmercury (MeHg) dynamics in mangrove sediments is important, as MeHg can potentially pose risks to marine biota and people that rely on these ecosystems. While the important role of sulfate-reducing bacteria in MeHg formation has been examined in this ecologically important habitat, the contribution of non-Hg methylating communities on MeHg production remains particularly unclear. Here, we collected sediment samples from 13 mangrove forests in south China and examined the geochemical parameters and microbial communities related to the Hg methylation. MeHg concentrations were significantly correlated to the OM-related parameters such as organic carbon content, total nitrogen, and dissolved organic carbon concentrations, suggesting the importance of OM in the MeHg production. Sulfate-reducing bacteria were the major Hg-methylators in mangrove sediments. Desulfobacteraceae and Desulfobulbaceae dominated the Hg-methylating microbes. Classification random forest analysis detected strong co-occurrence between Hg methylators and putative non-Hg methylators, thus suggesting that both types of microorganisms contribute to the MeHg dynamics in the sediments. Our study provides an overview of MeHg contamination in south China and advances our understanding of Hg methylation in mangrove ecosystems.

Biogeochemical controls on methylmercury distribution in a subtropical wetland ecosystem.

Wetlands are widely regarded as biogeochemical hotspots of mercury methylation but little is known regarding such roles of mangrove forests. Here, we examined the detailed depth profile of mercury, methylmercury, and organic matter in surface sediments within an estuarine pond at Mai Po Nature Reserve in Hong Kong, China. There is a progressive enrichment of organic matter in sites closer to mangrove forests, and methylmercury showed a significantly positive correlation with organic matter content (p < 0.001). Methylmercury in sediments is significantly (p < 0.05) higher in the summertime when the temperature is elevated but salinity is reduced. Further, sediments at or near the mangrove forest have lower carbon to nitrogen ratio, which may imply more labile organic matter in these organic-rich sediments that can promote microbial mercury methylation. In summary, mangrove forests can enhance net methylmercury production and increase the risk to the migratory birds overwintering in this internationally important wetland.

Soil Geobacteraceae are the key predictors of neurotoxic methylmercury bioaccumulation in rice.

Contamination of rice by the potent neurotoxin methylmercury (MeHg) originates from microbe-mediated Hg methylation in soils. However, the high diversity of Hg methylating microorganisms in soils hinders the prediction of MeHg formation and challenges the mitigation of MeHg bioaccumulation via regulating soil microbiomes. Here we explored the roles of various cropland microbial communities in MeHg formation in the potentials leading to MeHg accumulation in rice and reveal that Geobacteraceae are the key predictors of MeHg bioaccumulation in paddy soil systems. We characterized Hg methylating microorganisms from 67 cropland ecosystems across 3,600 latitudinal kilometres. The simulations of a rice-paddy biogeochemical model show that MeHg accumulation in rice is 1.3-1.7-fold more sensitive to changes in the relative abundance of Geobacteraceae compared to Hg input, which is recognized as the primary parameter in controlling MeHg exposure. These findings open up a window to predict MeHg formation and accumulation in human food webs, enabling more efficient mitigation of risks to human health through regulations of key soil microbiomes.

Methylmercury photodegradation in paddy water: An overlooked process mitigating methylmercury risks.

Photodegradation is critical to reduce the potent neurotoxic methylmercury (MeHg) in water and its subsequent accumulation along food chains. However, this process has been largely ignored in rice paddies, which are hotspots of MeHg production and receive about a quarter of the world's developed freshwater resources. Here, we reported that significant MeHg photodegradation, primarily mediated by hydroxyl radicals, occurs in the overlying water during rice growth. By incorporating field-measured light interception into a rice paddy biogeochemistry model, as well as photodegradation rates obtained from 42 paddy soils stretching ∼3500 km across China, we estimated that photodegradation reduced MeHg concentrations in paddy water and rice by 82 % and 11 %, respectively. Without photodegradation, paddy water could be a significant MeHg source for downstream ecosystems, with an annual export of 178 - 856 kg MeHg to downstream waters in China, the largest rice producer. These findings suggest that photodegradation in paddy water is critical for preventing greater quantities of MeHg entering human food webs.

Wildland-urban interface wildfire increases metal contributions to stormwater runoff in Paradise, California.

The 2018 Camp Fire was a large late-year (November) wildfire that produced an urban firestorm in the Town of Paradise, California, USA, and destroyed more than 18 000 structures. Runoff from burned wildland areas is known to contain ash, which can transport contaminants including metals into nearby watersheds. However, due to historically infrequent occurrences, the effect of wildland-urban interface (WUI) fires, such as the Camp Fire, on surface water quality has not been well-characterized. Therefore, this study investigated the effects of widespread urban burning on surface water quality in major watersheds of the Camp Fire area. Between November 2018 and May 2019, 140 surface water samples were collected, including baseflow and stormflow, from burned and unburned watersheds with varying extent of urban development. Samples were analyzed for total and filter-passing metals, dissolved organic carbon, major anions, and total suspended solids. Ash and debris from the Camp Fire contributed metals to downstream watersheds via runoff throughout the storm season. Increases in concentration up to 200-fold were found for metals Cr, Cu, Ni, Pb, and Zn in burned watersheds compared to pre-fire values. Total concentrations of Al, Cd, Cu, Pb, and Zn exceeded EPA aquatic habitat acute criteria by up to 16-fold for up to five months after the fire. To assess possible transport mechanisms and bioavailability, a subset of 18 samples was analyzed using four filters with nominal pore sizes ranging from 0.22 to 1.2 µm to determine the particulate size distribution of metals. Trace and major metals (Al, Ba, Co, Cr, Cu, Fe, Hg, Mn, Ni, Pb, and Zn) were found mostly associated with larger grain sizes (>0.45 µm), and some metals (Al, Cr, Fe, and Pb) also included a substantial colloidal phase (0.22 to 0.45 µm). This study suggests that fires in the wildland-urban interface increase metal concentrations, mainly through particulate driven transport. The metals with the largest increases are likely from anthropogenic disaster materials, though biomass ash also is a major contributor to water quality. The increase in metals following WUI burning may have adverse ecological impacts.

Tree foliage as a net accumulator of highly toxic methylmercury.

Tree canopies are known to elevate atmospheric inputs of both mercury (Hg) and methylmercury (MeHg). While foliar uptake of gaseous Hg is well documented, little is known regarding the temporal dynamics and origins of MeHg in tree foliage, which represents typically less than 1% of total Hg in foliage. In this work, we examined the foliar total Hg and MeHg content by following the growth of five individual trees of American Beech (Fagus grandifolia) for one growing season (April-November, 2017) in North Carolina, USA. We show that similar to other studies foliar Hg content increased almost linearly over time, with daily accumulation rates ranging from 0.123 to 0.161 ng/g/day. However, not all trees showed linear increases of foliar MeHg content along the growing season; we found that 2 out of 5 trees showed elevated foliar MeHg content at the initial phase of the growing season but their MeHg content declined through early summer. However, foliar MeHg content among all 5 trees showed eventual increases through the end of the growing season, proving that foliage is a net accumulator of MeHg while foliar gain of biomass did not "dilute" MeHg content.

Silica nanoparticle accumulation in plants: current state and future perspectives.

With their excellent biocompatibility, adjustable size, and high specific surface area, silica nanoparticles (SiO2 NPs) offer an alternative to traditional bulk fertilizers as a means to promote sustainable agriculture. SiO2 NPs have been shown to promote the growth of plants and to reduce the negative effects of biotic and abiotic stresses, but their bioaccumulation is a crucial factor that has been overlooked in studies of their biological effects. In this review, the techniques to quantify and visualize SiO2 NPs in plants were examined first. We then provide a summary of the current state of knowledge on the accumulation, translocation, and transformation of SiO2 NPs in plants and of the factors (e.g., the physicochemical properties of SiO2 NPs, plant species, application mode, and environmental conditions) that influence SiO2 NP bioaccumulation. The challenges in analyzing NP-plant interactions are considered as well. We conclude by identifying areas for further research that will advance our understanding of NP-plant interactions and thus contribute to more sustainable, eco-friendly, nano-enabled approaches to improving crop nutrient supplies. The information presented herein is important to improve the delivery efficiency of SiO2 NPs for precision and sustainable agriculture and to assess the safety of SiO2 NPs during their application in agriculture.

Micro(nano)plastic pollution in terrestrial ecosystem: emphasis on impacts of polystyrene on soil biota, plants, animals, and humans.

Pollution with emerging microscopic contaminants such as microplastics (MPs) and nanoplastics (NPs) including polystyrene (PS) in aquatic and terrestrial environments is increasingly recognized. PS is largely used in packaging materials and is dumped directly into the ecosystem. PS micro-nano-plastics (MNPs) can be potentially bioaccumulated in the food chain and can cause human health concerns through food consumption. Earlier MP research has focused on the aquatic environments, but recent researches show significant MP and NP contamination in the terrestrial environments especially agricultural fields. Though PS is the hotspot of MPs research, however, to our knowledge, this systematic review represents the first of its kind that specifically focused on PS contamination in agricultural soils, covering sources, effects, and ways of PS mitigation. The paper also provides updated information on the effects of PS on soil organisms, its uptake by plants, and effects on higher animals as well as human beings. Directions for future research are also proposed to increase our understanding of the environmental contamination of PS in terrestrial environments.

Declines of methylmercury along a salinity gradient in a low-lying coastal wetland ecosystem at South Carolina, USA.

Wetlands are widely regarded as biogeochemical hotspots of highly toxic methylmercury (MeHg), mainly mediated by sulfate-reducing bacteria. In low-lying coastal wetlands, sea level rise, a phenomenon caused by global climate change, is slowly degrading numerous healthy freshwater forested wetlands into salt-degraded counterparts with a nickname "ghost forests", and eventually converting them to saltmarshes. However, little is known about the changes of mercury (Hg) methylation, bioaccumulation, and biomagnification along the forest-to-saltmarsh gradient. Here, we conducted extensive field sampling in three wetland states (healthy forested wetlands, salt-degraded forested wetlands, and saltmarsh) along a salinity gradient (from 0 to 9.4 ppt) in Winyah Bay, South Carolina, USA. We found that in our study wetland systems the saltmarshes had the lowest levels of both total Hg and MeHg in sediments and biota, as compared to healthy forested wetlands and saltwater-degraded ghost forests. Our results suggest that the slow conversion of forested wetland to saltmarsh could reduce net MeHg production in our study wetland systems, which we hypothesized that could be attributed to increased sulfate reduction and excessive buildup of sulfide in sediment that inhibits microbial Hg methylation, and/or reduced canopy density and increased photodegradation of MeHg. However, it should be noted that biogeochemical MeHg responses to salinity changes may be site-specific and we urge more similar studies in other wetland systems along a salinity gradient. Therefore, long-term salinization of coastal wetlands and the slow conversion of forests to marshes could decrease long-term exposure of toxic MeHg levels in coastal food webs that are similar to our system, and ultimately reduce human exposure to this neurotoxin.

Burn Intensity Drives the Alteration of Phenolic Lignin to (Poly) Aromatic Hydrocarbons as Revealed by Pyrolysis Gas Chromatography-Mass Spectrometry (Py-GC/MS).

High-intensity wildfires alter the chemical composition of organic matter, which is expected to be distinctly different from low-intensity prescribed fires. Herein, we used pyrolysis gas chromatography/mass spectrometry (Py-GC/MS), in conjunction with solid-state 13C nuclear magnetic resonance (NMR) and Fourier transform infrared (FT-IR) spectroscopy, to assess chemical alterations from three wildfires and a long-term frequent prescribed fire site. Our results showed that black ash formed under moderate intensity burns contained less aromatic (ArH), polyaromatic hydrocarbon (PAH), and nitrogen-containing compounds (Ntg) but more lignin (LgC) and phenol compounds (PhC), compared to white ash formed under high intensity burns. Both 13C NMR and FT-IR confirmed a higher relative percentage of carboxyl carbon in white ash, indicating the potential for higher water solubility and more mobile carbon, relative to black ash. Compared to wildfires, ash from low-intensity prescribed fire contained less ArH, PAH, and Ntg and more LgC and PhC. Controlled laboratory burning trials indicated that organic matter alteration was sensitive to the burn temperature, but not related to the fuel type (pine vs fir) nor oxygen absence/presence at high burn temperatures. This study concludes that higher burn temperatures resulted in higher (poly)aromatic carbon/nitrogen and lower lignin/phenol compounds.

Ecological Traits of Fish for Mercury Biomonitoring: Insights from Compound-Specific Nitrogen and Stable Mercury Isotopes.

We coupled compound-specific isotopic analyses of nitrogen (N) in amino acids (δ15NGlu, δ15NPhe) and mercury stable isotopes (δ202Hg, Δ199Hg) to quantify ecological traits governing the concentration, variability, and source of Hg in largemouth bass (LB) and pike gudgeon (PG) across four rivers, South Korea. PG displayed uniform Hg concentration (56-137 ng/g), trophic position (TPcorrected; 2.6-3.0, n = 9), and N isotopes in the source amino acid (δ15NPhe; 7-13‰), consistent with their specialist feeding on benthic insects. LB showed wide ranges in Hg concentration (45-693 ng/g), TPcorrected (2.8-3.8, n = 14), and δ15NPhe (1.3-16‰), reflecting their opportunistic feeding behavior. Hg sources assessed using Hg isotopes reveal low and uniform Δ199Hg in PG (0.20-0.49‰), similar to Δ199Hg reported in sediments. LB displayed site-specific δ202Hg (-0.61 to -0.04‰) and Δ199Hg (0.53-1.09‰). At the Yeongsan River, LB displayed elevated Δ199Hg and low δ15NPhe, consistent with Hg and N sourced from the atmosphere. LB at the Geum River displayed low Δ199Hg and high δ15NPhe, both similar to the isotope values of anthropogenic sources. Our results suggest that a specialist fish (PG) with consistent ecological traits and Hg concentration is an effective bioindicator species for Hg. When accounting for Hg sources, however, LB better captures site-specific Hg sources.

Impacts of Forest Fire Ash on Aquatic Mercury Cycling.

Mercury (Hg) is a ubiquitous contaminant in the environment and its methylated form, methylmercury (MeHg), poses a worldwide health concern for humans and wildlife, primarily through fish consumption. Global production of forest fire ash, derived from wildfires and prescribed burns, is rapidly increasing due to a warming climate, but their interactions with aqueous and sedimentary Hg are poorly understood. Herein, we compared the differences of wildfire ash with activated carbon and biochar on the sorption of aqueous inorganic Hg and sedimentary Hg methylation. Sorption of aqueous inorganic Hg was greatest for wildfire ash materials (up to 0.21 µg g-1 or 2.2 µg g-1 C) among all of the solid sorbents evaluated. A similar Hg adsorption mechanism for activated carbon, biochar made of walnut, and wildfire ash was found that involves the formation of complexes between Hg and oxygen-containing functional groups, especially the -COO group. Notably, increasing dissolved organic matter from 2.4 to 70 mg C L-1 remarkably reduced Hg sorption (up to 40% reduction) and increased the time required to reach Hg-sorbent pseudo-equilibrium. Surprisingly, biochar and wildfire ash, but not activated carbon, stimulated MeHg production during anoxic sediment incubation, possibly due to the release of labile organic matter. Overall, our study indicates that while wildfire ash can sequester aqueous Hg, the leaching of its labile organic matter may promote production of toxic MeHg in anoxic sediments, which has an important implication for potential MeHg contamination in downstream aquatic ecosystems after wildfires.

Internal Dynamics and Metabolism of Mercury in Biota: A Review of Insights from Mercury Stable Isotopes.

Monitoring mercury (Hg) levels in biota is considered an important objective for the effectiveness evaluation of the Minamata Convention. While many studies have characterized Hg levels in organisms at multiple spatiotemporal scales, concentration analyses alone often cannot provide sufficient information on the Hg exposure sources and internal processes occurring within biota. Here, we review the decadal scientific progress of using Hg isotopes to understand internal processes that modify the speciation, transport, and fate of Hg within biota. Mercury stable isotopes have emerged as a powerful tool for assessing Hg sources and biogeochemical processes in natural environments. A better understanding of the tissue location and internal mechanisms leading to Hg isotope change is key to assessing its use for biomonitoring. We synthesize the current understanding and uncertainties of internal processes leading to Hg isotope fractionation in a variety of biota, in a sequence of better to less studied organisms (i.e., birds, marine mammals, humans, fish, plankton, and invertebrates). This review discusses the opportunities and challenges of using certain forms of biota for Hg source monitoring and the need to further elucidate the physiological mechanisms that control the accumulation, distribution, and toxicity of Hg in biota by coupling new techniques with Hg stable isotopes.

Effects and mechanisms of organic matter regulating the methylmercury dynamics in mangrove sediments.

Mangrove ecosystems serve as an important carbon sink but also could be a hotspot that produces neurotoxic methylmercury (MeHg). Although many studies have focused on mercury (Hg) contamination in this carbon-rich ecosystem, our understanding of the effects and mechanisms of the organic matter (OM) regulation of MeHg production in mangrove sediments is still limited. Here, we examined the effects of Hg contamination and OM enrichment on MeHg production in anoxic mangrove sediments and identified the major microbial guilds attending this process. The mangrove sediments possessed a high potential for producing MeHg, but this was counterbalanced by its rapid degradation. Sulfate-reducing bacteria (SRB) such as Desulfobacterales, Desulfovibrionales, and Syntrophobacterales were the major methylators. OM diagenesis significantly changed the biogeochemical conditions, accelerating MeHg degradation in the sediments. The enhanced MeHg degradation could be attributed to the abundant sulfide produced during OM decomposition, which could potentially inhibit the Hg methylation by immobilization of inorganic Hg, abiotically degrade MeHg, and favor the non-mer-mediated degradation of MeHg by SRB. Our study provides both geochemical and microbial clues that can partly explain the low MeHg levels widely observed in mangrove sediments.

Alteration of acute toxicity of inorganic and methyl mercury to Daphnia magna by dietary addition.

Acute toxicity of inorganic mercury [Hg(II)] and methylmercury (MeHg) to Daphnia magna was characterized using a 48-h static, non-renewal acute toxicity test, in which we compared the toxicity of Hg(II) and MeHg in the absence (water-only) and presence of diet [green alga (Raphidocelis subcapitata), yeast, Cerophyll, and trout chow (YCT), or both]. Overall, Hg(II) is more toxic to D. magna than MeHg, with 48-h median lethal concentrations (LC50s) being 4.3 µg/L (95% confidence interval: 4.1-4.5 µg/L) for Hg(II) and 14.3 µg/L (13.2-15.3 µg/L) for MeHg. For Hg(II), the addition of any diet would significantly increase its 48-h LC50, but the 48-h LC50 for MeHg decreased significantly to 7.1 µg/L (6.4-7.8 µg/L) with the algal addition. We also show that the addition of diets significantly influenced the levels and speciation (dissolved vs. particulate) of both Hg forms in the test solution. The bioaccumulation of Hg(II) and MeHg was impacted by the dietary addition, and it appears that the body residue level triggering mortality varied widely among treatments. The results suggest that standard short-term toxicity tests (water-only) should be supplemented with extra tests with dietary addition to provide a more environmentally relevant estimation of short-term toxicity of chemical compounds.

Microplastics interaction with terrestrial plants and their impacts on agriculture.

Microplastics (MPs) are widespread in natural ecosystems and have attracted considerable attention from scientists all over the world because they are believed to threaten every life form. In addition to their potential physical and chemical effects on organisms, MPs may act as a carrier for many micropollutants, including antibiotics, heavy metals, and others. Over the last 10-15 yr, extensive research has been carried out on MPs in marine environments regarding their sources, fate, and toxicity. However, studies concerning their accumulation in the soil ecosystem, uptake, internalization, and impacts on photosynthetic components of the terrestrial ecosystem and risk assessments have been scanty. Thus, there is a large knowledge gap on the extent to which terrestrial environments, especially agroecosystems, are affected by MPs and their subsequent risks to human health. This review summarizes up-to-date findings about MPs on terrestrial environments and provides guidelines for future studies regarding the phytotoxic effects of MPs on plants; the mechanism of uptake and translocation in plant tissues; detection tools for MPs in plants; impacts on plant growth, plant development, and agricultural productivity; and, most important, the future prospects of MPs interaction and accumulation in plants.

Methyl mercury triggers endothelial leukocyte adhesion and increases expression of cell adhesion molecules and chemokines.

Cardiovascular disease is the leading cause of morbidity, mortality, and health care costs in the USA, and around the world. Among the various risk factors of cardiovascular disease, environmental and dietary exposures to methyl mercury, a highly toxic metal traditionally labeled as a neurotoxin, have been epidemiologically linked to human cardiovascular disease development. However, its role in development and promotion of atherosclerosis, an initial step in more immediately life-threatening cardiovascular diseases, remains unclear. This study was conducted to examine the role that methyl mercury plays in the adhesion of monocytes to human microvascular endothelial cells (HMEC-1), and the underlying mechanisms. Methyl mercury treatment significantly induced the adhesion of monocyte to HMEC-1 endothelial cells, a critical step in atherosclerosis, while also upregulating the expression of proinflammatory cytokines interleukin-6, interleukin-8. Further, methyl mercury treatment also upregulated the chemotactic cytokine monocyte chemoattractant protein-1 and intercellular adhesion molecule-1. These molecules are imperative for the firm adhesion of leukocytes to endothelial cells. Additionally, our results further demonstrated that methyl mercury stimulated a significant increase in NF-κB activation. These findings suggest that NF-κB signaling pathway activation by methyl mercury is an important factor in the binding of monocytes to endothelial cells. Finally, by using flow cytometric analysis, methyl mercury treatment caused a significant increase in necrotic cell death only at higher concentrations without initiating apoptosis. This study provides new insights into the molecular actions of methyl mercury that can lead to endothelial dysfunction, inflammation, and subsequent atherosclerotic development.

Feasibility of Sijunzi Tang (Chinese medicine) to enhance protein disulfide isomerase activities for reactivating malate dehydrogenase deactivated by polycyclic aromatic hydrocarbons.

The objective of this research is to investigate the enzymatic activities between protein disulfide isomerase (PDI) found in animals and plants and the properties found in a commonly used Chinese medicine called Sijunzi Tang. During the investigation, PDI, which is a monomer with a molecular mass of 57.0 kDa, was used to reactivate malate dehydrogenase (MDH). However, with the interference of polycyclic aromatic hydrocarbons (PAHs), evidence indicates that such chemicals are carcinogenic, mutagenic, and toxic to humans. The enzymatic activity of PDI found in animal's liver and plant was 1657 folds of purification; 0.284 unit/mg of enzyme activity, and 5694.4 folds of purification; 1.00 unit/mg of enzyme activity, respectively. PDI extracted in treated animal and plant tissue revealed 2.40% and 80.44% of regaining MDH enzymatic activity, respectively. Although in its initial phase of investigation, it is assumed that the properties found in Sijunzi Tang can help regain enzymatic activity in those affected by xenobiotic substances, thus, making it a potential ingredient in assisting with PDI functions.

Examination of mercury contamination from a recent coal ash spill into the Dan River, North Carolina, United States.

Coal ash spills occasionally occur due to the accidental failure of surface impoundments, and toxic metal-laden ash can pose a serious health threat to adjacent aquatic ecosystems. Here, we performed an investigation into longitudinal variations of mercury (Hg) contamination in the Dan River (North Carolina, United States) about 17 and 29 months after a February 2014 coal ash spill incident, in which the reported Hg concentrations in the spilled coal ash (210 ng/g) were 1-2 orders of magnitude higher than the river sediments (2-61 ng/g). We examined total Hg (THg) and methyl Hg (MeHg) in sediments from 0 to 65 km downstream of the spill, and found that most of the variations of THg and MeHg in surface sediments (0-16 cm) could be well accounted by the organic matter content and appeared to be not contaminated by Hg derived from coal ash. In examining MeHg bioaccumulation in invertebrates (aquatic and riparian) and fish in the Dan River and fish in a reservoir downstream of Dan River, we found no evidence of elevated MeHg bioaccumulation due to the 2014 coal ash spill. Thus, we concluded that Hg contamination from the coal ash spill is largely absent in the Dan River for both surface sediments and biota within the first three years of spill (until 2017), even though the majority of coal ash may be buried deeper in the sediment in the river channel and/or the downstream reservoir. Alternatively, the Hg associated with the coal ash is largely not bioavailable for extensive microbial Hg methylation. The findings provide useful insights into remediation strategies for this incident and other coal ash spills.